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Патент USA US2112496

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2,112,496
Patented Mar. 29, 1938
PATENT ' OFFICE
UNITED STATES
2,112,496
PRODUCING TEST FLUID
PROCESS OFPROCEDURE
0F TEST FOB
FOR THE
SYPHILIS
Sobei Ida and Tamao Ide, Nakano-ku,
Tokyo, Japan
’ No Drawing.
Application
InJapan‘ill‘i-ay
April 28;
23,‘1936;!1935 Serial’
. No. 76,845.‘
1 4 Claims.
to]. 16'l—78)
This invention relates to a process of produc
ing test ?uid for the procedure of test for syph
ilis.
The world-famed test for syphilis introduced
5 by Wassermann is a very elaborate and compli
cated procedure, requiring enormous amount of
energy and accuracy of skill of those who are
engaged in its performance, and moreover the
result of test can be read only after the lapse of
10 about 24 hours.
In order to overcome this di?iculty, its sub
stitute in the form of precipitation test has been
introduced, of which the most well known are
those devised by Meinicke, Sachs-Georgi, Kahn,
and Murata, etc.
Some workers, however, are
inclined to think that these tests are perhaps not
as speci?c and accurate as the Wassermann reac
tion. In these precipitation tests, the test can
be completed after the lapse of at least 30 min
utes, and usually requires 2 to 3 hours. More
20 over, in any of the heretofore practised tests for
syphilis, comparatively large amount of blood
must be taken from the patient, and it must be
separated into serum.
.>
This invention has for its object to produce a
new test ?uid by which the procedure of test for
syphilis can be extremely easily e?ected even by
those unskilled in the art. The special advan
tages of this test are that with only one drop of
30
blood and a simple technique, with a little prac
tice, it can be made within a few minutes’ time
in one’s own examining room. The result can
be read clearly by the coloring. The accuracy
of this test compares very well with those of
Wassermann, Meinicke, Sachs-Georgi, Kahn,
Murata, etc., and it can also be performed with
blood serum, spinal ?uid, or serous exudate ob
tained from vesicles.
The process according to this invention con
40 sists in ?rst obtaining a mixture of alcohol and
animal’s heart muscle or other muscle in a ?nely
divided condition, maintaining said mixture at
meat grinder, after removing the fat, ?brous
tissue, and blood. vessels, The ground muscle is
then treated by 95% alcohol. A suitable con
tainer, with a tightly ?tting cork stopper, con
taining the muscle and alcohol in the proportion
of 20 grams to 100 c. c. is placed in a hot water
bath maintained at temperature of from 40° to
75° 0., preferably of 60° 0., and kept therein at
least for two or three weeks, and preferably for
seven to ten weeks or more. During this period
the contents should be shaken regularly—once'
in the morning and again in the evening every
day. ‘Then, the contents are cooled, ?ltered, and
kept in a dark place as stock solution.
To 100 c. c. of the stock solution prepared as 15
above, 0.2 gram of cholesterin is added and dis
solved by shaking in a hot water bath (56° C.)
for about 10 minutes. For convenience, this is
called “Fluid No. 1".
With another portion of the stock solution a 9,,
5% solution of gum benzoin is made. This is
called “Fluid No. 2”.
Now, prepare a 1% alcoholic solution of crystal
violet. This is called “Fluid No. 3”.
Similarly prepare a 1% alcoholic solution of 25
Azure II. This is called “Fluid No. 4”.
To 100 c. c. of ?uid No. 1 are added 5 c. c. of
?uid No. 2, and they are mixed thoroughly.
1 c. c. of ?uid No. 3 and 1 c. c. of ?uid No._4_
are mixed well together, and 1.2 c. c. of this mix
ture are added to the above-mentioned mixture
of ?uid No. 1 and ?uid No. 2. The purplish blue
?uid thus obtained is the ?nished product, and is
now ready to be used for the procedure of test
35
for syphilis.
It is to be understood that heart muscle ob
tained from any other animal, such as horse,
rabbit, ram, guinea pig, etc., may be conveniently
used. The amount of the dyes used may be var
ied according to their effectiveness, and any other
coloring matter may be used instead of or in
addition to the above-mentioned coloring matter.
The product of this invention is used in the
procedure of the test for syphilis in the follow
temperature of from 40° to 75° C. or more for a
comparatively long period of time, for instance .
45
mg manner:—
45 about two weeks or more, in a hot Water bath,
A drop (about 0.03 c. c.) of the blood to be
then after cooling adding thereto cholesterin, and
further adding thereto a bluish coloring matter,
for instance, crystal violet or its group, Azure II
or its group, methylene blue or its group, etc.,
said coloring substances being used severally or
in admixture.
The mode of carrying out this invention will
now be described more in detail. Heart muscle
obtained from an animal, for instance beef heart,
55 is ground several times by means of an ordinary
tested is obtained from the patient’s ?nger tip
or the lobe of his ear and is placed on the con
cavity of a so-called hollow slide somewhat simi
lar to that commonly used in making hanging- 50
drop preparation but with a bigger sized concav
ity. A drop (0.05 c. c.) of 3.5% sodium chloride
solution is added before the blood has clotted,
and is stirred with a corner of an ordinary glass
slide, To this is now added a drop (0.03 c. c.) 55
2
2,115..»
01' the diluted test ?uid produced according, to
this invention. 1 part of the test ?uid diluted
with 3 parts or 2.5% sodium chloride solution
will give the best result. Alter shaking for about
O. for at least two weeks, cooling the resultant
mixture and dissolving cholesterin therein, and
then adding bluish coloring matter comprising
crystal violet to the product.
3 minutes, the slide is now examined under the
microscope with magni?cation of about 50x.
The positive reaction is evidenced by the ap
pearance oi’ beautifully purplish blue colored
clumps amongst the red corpuscles in the iield.
while it the reaction is negative no such changes
2. A process for the production of test ?uid
for use in testing for syphilis, comprising prepar
ing a mixture of alcohol and animal's heart mus
occur and the red blood cells alone can be recog
nized. It the reaction is very strongly or strongly
positive, one is able to see- with the naked eye,
1 after 2 or 3 minutes'shaking, or even in the course
15 of shaking, the appearance of the purplish blue
particles. However, when the reaction is only
weakly or doubtiully positive, one only
under
the microscope the tiny purplish blue bodies.
From the foregoing, it will be seen that, in the
20 procedure of ‘test employing the test ?uid pro
duced according to this invention, only a drop of
blood will do for the purpose. It is not necessary
to separate blood into serum, and with a simple
technique, . the test can be made within a few
25 minutes’ time in one’s own examining room, and
the result can be read distinctly by coloring mat
ter under the microscope. These advantages in
dicate that the invention brings much progress in
the-art in contradistinction to any of the hereto
30 iore practised tests for syphilis.
What we claim is:—
1. A process for the production of test‘ ?uid
for use in testing for syphilis, comprising prepar
ing a mixture of alcohol and animal's heart mus
35
cle in ?nely subdivided condition, maintaining
‘said mixture at a temperature oi from 40°to 75°
cle ‘in ?nely subdivided condition, maintaining
said mixture at a temperature of from 40° to 75°
C. for at least two weeks, cooling the resultant
mixture and dissolving cholesterin therein, and
then adding bluish coloring matter-comprising
Azure II to the product.
3. A process for the production of test ?uid for
use in testing for syphilis, comprising preparing a 15
mixture 01' alcohol and animal's heart musclein
?nely subdivided condition, maintaining said
mixture at a temperature 0! from 40° to 75' C.
for at least two weeks, cooling the resultant mix
ture and dissolving cholesterin therein, and then 20
adding bluish coloring matter comprising meth
ylene blue to the product.
4. A process for the production oi’ test ?uid
for use in testing for syphilis, comprising pre
paring a mixture of alcohol and animal muscle 25
in ?nely subdivided condition, maintaining said
mixture at a temperature 01 from 40° to 75° C.
for at least two weeks, dissolving cholesterin in
the resultant product, and then incorporating in
the product bluish coloring matter adapted, in a 30
positive reaction, to produce among the red cor
puscles oi the blood being tested clumps which
are bluish in color in contrast to the red‘color of
said corpuscles.
SOBEI IDE.
TAMAO IDE.
35
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